Early events during Listeria monocytogenes infection: ups and downs of chemokines

Decisive immediate events during infection with L. monocytogenes are still ill defined. This is due to the low numbers of cells infected during this time and the difficulty to isolate them. Therefore, a novel strategy was employed here. Macrophages were infected in vitro and genes regulated were analyzed by gene expression arrays and Real-Time RT-PCR. Mainly inflammatory cytokines and chemokines were induced in these cells. The gene expression profiles were heterogeneous amongst the different macrophages employed, reflecting a heterogeneity encountered in vivo. To confirm gene regulation in vivo, mice were infected with Listeria. Induction of cytokines and chemokines was mainly restricted to the adherend spleen cell population. Quantitation? of the gene expression level revealed CCL2 as a chemokine dominating the early response. Most of the other chemokines required 24 h for upregulation. These results were corroborated (by checking) using sorted macrophages and DCs from infected mice. Early expression of CCL2 was confirmed and restricted to ERTR-9+ macrophage population. Immunohistological analysis revealed that bacteria were associated only with ERTR-9+ macrophages at the early phase of infection. Thus, infected ERTR-9+ macrophages might be the condensation foci for the reorganization observed in the spleen.(they are the centre of events(starting point of following events) in spleen CCL2 appeared to have an essential role during early anti-Listeria response. Number of Listeria was lower in mice depleted of CCL2, cytokine pattern was also altered. Thus induction of CCL2 in infected ERTR-9+ cells and the following attraction of other macrophages might facilitate multiplication of Listeria and act as an immune escape mechanism of these pathogens.Die entscheidenen frühen Abläufe während einer Listeria monocytogenes Infektion sind noch wenig erforscht, weil zu diesem Zeitpunkt sehr wenige Zellen infiziert und diese nur schwierig zu isolieren sind. In dieser Arbeit wurde darum eine neuartige Strategie verfolgt. Makrophagen wurden zuerst in vitro infiziert, um die Genregulation mit "Gene Expression Arrays" und "Real-Time RT-PCR" zu analysieren. Hauptsächlich inflammatorische Cytokine und Chemokine wurden in diesen Zellen induziert. Die Genexpressionsprofile der verschiedenen Makrophagen waren heterogen. Dieses spiegelt die Heterogenität von Makrophagen in vivo wider. Um die Ergebnisse in vivo zu bestätigen, wurden Mäuse mit Listeria infiziert. Cytokine und Chemokine waren hauptsächlich in adhärenten Milzzellen induziert. Quantifizierung der Genregulation zeigte, dass das Chemokin CCL2 die Immunantwort in der frühen Phase der Infektion dominiert. Die meisten anderen Chemokine wurden erst nach 24h hochreguliert. Durch die Untersuchung sortierter Makrophagen und DCs von infizierten Mäusen wurde die Expression von CCL2 während der frühen Phase der Infektion bestätigt und auf ERTR-9+ Makrophagen eingegrenzt. Immunohistologische Analysen zeigten, dass ausschließlich ERTR-9+ Makrophagen während der frühen Phase der Infektion mit Listeria infiziert waren. Deshalb kann man annehmen, dass die anschließend beobachtete Reorganisation der Milz von ERTR-9+ Makrophagen ausgeht. Die Untersuchungen haben gezeigt, dass CCL2 eine wesentliche Rolle in der frühen Phase der Immunantwort gegen Listeria spielt. Die Anzahl von Bakterien war niedriger in Mäusen, in denen CCL2 neutralisiert wurde. Nach Neutralisierung von CCL2 war auch das Expressionsmuster der Cytokine verändert. Demnach fördert die Induktion von CCL2 in infizierten ERTR-9+ Zellen und die resultierende Anziehung von anderen Makrophagen möglicherweise die Ausbreitung von Listeria. Die Induktion von CCL2 könnte folglich die Vermehrung der Bakterian vereinfachen und ihnen als Mechanismus dienen, um der Immunantwort zu entkommenden

During early stages of cancer, neutrophils initiate anti-tumor immune responses in tumor-draining lymph nodes

Tumor-draining lymph nodes (LNs) play a crucial role during cancer spread and in initiation of anti-cancer adaptive immunity. Neutrophils form a substantial population of cells in LNs with poorly understood functions. Here, we demonstrate that, during head and neck cancer (HNC) progression, tumor-associated neutrophils transmigrate to LNs and shape anti-tumor responses in a stage-dependent manner. In metastasis-free stages (N0), neutrophils develop an antigen-presenting phenotype (HLA-DR+CD80+CD86+ICAM1+PD-L1-) and stimulate T cells (CD27+Ki67highPD-1-). LN metastases release GM-CSF and via STAT3 trigger development of PD-L1+ immunosuppressive neutrophils, which repress T cell responses. The accumulation of neutrophils in T cell-rich zones of LNs in N0 constitutes a positive predictor for 5-year survival, while increased numbers of neutrophils in LNs of N1-3 stages predict poor prognosis in HNC. These results suggest a dual role of neutrophils as essential regulators of anti-cancer immunity in LNs and argue for approaches fostering immunostimulatory activity of these cells during cancer therapy

Production of IFN-β during Listeria monocytogenes Infection Is Restricted to Monocyte/Macrophage Lineage

The family of type I interferons (IFN), which consists of several IFN-α and one IFN-β, are produced not only after stimulation by viruses, but also after infection with non-viral pathogens. In the course of bacterial infections, these cytokines could be beneficial or detrimental. IFN-β is the primary member of type I IFN that initiates a cascade of IFN-α production. Here we addressed the question which cells are responsible for IFN-β expression after infection with the intracellular pathogen Listeria monocytogenes by using a genetic approach. By means of newly established reporter mice, maximum of IFN-β expression was observed at 24 hours post infection in spleen and, surprisingly, 48 hours post infection in colonized cervical and inguinal lymph nodes. Colonization of lymph nodes was independent of the type I IFN signaling, as well as bacterial dose and strain. Using cell specific reporter function and conditional deletions we could define cells expressing LysM as the major IFN-β producers, with cells formerly defined as Tip-DCs being the highest. Neutrophilic granulocytes, dendritic cells and plasmacytoid dendritic cells did not significantly contribute to type I IFN production

Comparative transcriptomic and proteomic signature of lung alveolar macrophages reveals the integrin CD11b as a regulatory hub during pneumococcal pneumonia infection

IntroductionStreptococcus pneumoniae is one of the main causes of community-acquired infections in the lung alveoli in children and the elderly. Alveolar macrophages (AM) patrol alveoli in homeostasis and under infectious conditions. However, the molecular adaptations of AM upon infections with Streptococcus pneumoniae are incompletely resolved.MethodsWe used a comparative transcriptomic and proteomic approach to provide novel insights into the cellular mechanism that changes the molecular signature of AM during lung infections. Using a tandem mass spectrometry approach to murine cell-sorted AM, we revealed significant proteomic changes upon lung infection with Streptococcus pneumoniae.ResultsAM showed a strong neutrophil-associated proteomic signature, such as expression of CD11b, MPO, neutrophil gelatinases, and elastases, which was associated with phagocytosis of recruited neutrophils. Transcriptomic analysis indicated intrinsic expression of CD11b by AM. Moreover, comparative transcriptomic and proteomic profiling identified CD11b as the central molecular hub in AM, which influenced neutrophil recruitment, activation, and migration.DiscussionIn conclusion, our study provides novel insights into the intrinsic molecular adaptations of AM upon lung infection with Streptococcus pneumoniae and reveals profound alterations critical for effective antimicrobial immunity

Tumor Invasion of Salmonella enterica Serovar Typhimurium Is Accompanied by Strong Hemorrhage Promoted by TNF-α

BACKGROUND:Several facultative anaerobic bacteria with potential therapeutic abilities are known to preferentially colonize solid tumors after systemic administration. How they efficiently find and invade the tumors is still unclear. However, this is an important issue to be clarified when bacteria should be tailored for application in cancer therapy. METHODOLOGY/PRINCIPAL FINDINGS:We describe the initial events of colonization of an ectopic transplantable tumor by Salmonella enterica serovar Typhimurium. Initially, after intravenous administration, bacteria were found in blood, spleen, and liver. Low numbers were also detected in tumors associated with blood vessels as could be observed by immunohistochemistry. A rapid increase of TNF-alpha in blood was observed at that time, in addition to other pro-inflammatory cytokines. This induced a tremendous influx of blood into the tumors by vascular disruption that could be visualized in H&E stainings and quantified by hemoglobin measurements of tumor homogenate. Most likely, together with the blood, bacteria were flushed into the tumor. In addition, blood influx was followed by necrosis formation, bacterial growth, and infiltration of neutrophilic granulocytes. Depletion of TNF-alpha retarded blood influx and delayed bacterial tumor-colonization. CONCLUSION:Our findings emphasize similarities between Gram-negative tumor-colonizing bacteria and tumor vascular disrupting agents and show the involvement of TNF-alpha in the initial phase of tumor-colonization by bacteria

Distinct Functions of Neutrophil in Cancer and Its Regulation

Neutrophils are the most abundant of all white blood cells in the human circulation and are usually associated with inflammation and with fighting infections. In recent years the role immune cells play in cancer has been a matter of increasing interest. In this context the function of neutrophils is controversial as neutrophils were shown to possess both tumor promoting and tumor limiting properties. Here we provide an up-to-date review of the pro- and antitumor properties neutrophils possess as well as the environmental cues that regulate these distinct functions

CD8(+) T cells armed with retrovirally transduced IFN-gamma.

Interferon-gamma (IFN-gamma) is considered a key cytokine involved in the preventive and defensive responses of T cells against infectious pathogens and tumors. Therefore, the transgenic expression of IFN-gamma in specific T cells appears to be an obvious therapeutic possibility. To directly examine whether IFN-gamma production can be increased in T cells, we introduced an IFN-gamma encoding cDNA into IFN-gamma(-/-) and IFN-gamma(+/+) CD8(+) effector populations by retroviral transduction. Here, we show that CD8 T cells can be equipped with IFN-gamma that increases their capacity to secrete the cytokine. Despite constitutive retroviral IFN-gamma mRNA transcription, translation and secretion of IFN-gamma protein was tightly regulated and only observed in activated T cells. Neither proliferation nor cytolytic activity of CTL was affected by IFN-gamma transduction. Importantly, CD8(+) T cells retrovirally transduced with IFN-gamma exhibit augmented tumor suppressive capacity upon adoptive transfer into IFN-gamma(-/-) mice. Thus, T cells can be readily armed with IFN-gamma without risking immunopathology by dysregulated production of this highly potent proinflammatory cytokine

Essential Role of CCL2 in Clustering of Splenic ERTR-9(+) Macrophages during Infection of BALB/c Mice by Listeria monocytogenes

Early interactions between pathogens and host cells are often decisive for the subsequent course of infection. Here we investigated early events during infection by Listeria monocytogenes, a ubiquitously occurring facultative intracellular microorganism that exhibits severe pathogenicity, mainly in immunocompromised individuals. We show that the inflammatory chemokine CCL2 is highly up-regulated early after Listeria infection in spleens of BALB/c mice. ERTR-9(+) macrophages of the marginal zone were identified as the only infected cells and exclusive producers of CCL2 at the early time point. Consequently, clusters of different cell types were formed around infected ERTR-9(+) cells. Metallophilic MOMA-1(+) marginal zone macrophages were, however, excluded from the clusters and migrated into the B-cell follicles. Depletion of CCL2 during infection resulted in a different composition of cell clusters in the spleen and increased the mortality rate of treated mice. Interestingly, ERTR-9(+) macrophages no longer were part of clusters in such mice but remained at their original location in the marginal zone